In March 1997, a new rhabdovirus was isolated from moribund cultured Japanese flounder Paralichthys olivaceus in sea water tank and cage culture systems in Kyung-Nam and Chun-Nam province, Korea. At temperature $15^{\circ}C$ the virus replicated and induced cytopathic effects (CPE), which progressed to eventual cytolysis, in susceptible cell lines, including RTG-2 and EPC. The CHES-214 cell line was refractory. Virus particles were bullet-shaped and measured $70nm{\times}100$ to 150 nm in size. The isolate was sensitive to pH 3, to diethyl ether, and to heat ($50^{\circ}C$ 5 min, $60^{\circ}C$ 1 min). Viral replication was not inhibited by $10^{-4}$ M 5-iododeoxyuridine. Virus infectivity was reduced by anti-HRV (8401-H) rabbit serum, but can not reduced by antisera against infectious hematopoietic necrosis virus (IHNV), chum salmon reovirus (CSV), retrovirus of salmonid (RVS) and infectious pancreatic necrosis virus (IPNV). HRV virus antigen was detected by fluorescent antibody test (FAT) in the cytoplasm of infected EPC cell. Purified isolates virions were composed of: polymerase (L), glycoprotein (G), nucleoprotein (N) and 2 matrix proteins (M1 and M2). Based upon their relative mobilities, the estimated molecular weights of the proteins were: L, 160 kDa; G, 55 kDa; N, 45 kDa; M1, 26 kDa; and M2, 22 kDa.
Nguyen, Ngoc Tuan;Lee, Kyoung;Kang, Ju Beom;Huang, Shir-Ly
Korean Journal of Microbiology
/
v.50
no.3
/
pp.210-215
/
2014
Pseudomonas nitroreducens TX1 was isolated from a rice field drainage in Taiwan. The bacterium is of special interest because of its capability to use a group of nonionic surfactants such as alkylphenol polyethoxylates even at high concentrations as a sole carbon source. In this study, a small cryptic circular plasmid, pTX1, was characterized from P. nitroreducens TX1. It is 2,286 bp in length with a GC content of 63.3% and harbors three open reading frames, $Rep_{pTX1}$ and functionally unidentified ORF1 and ORF2. The predicted $rep_{pTX1}$ gene product is homologous to Rep proteins of plasmids belonging to the pC194/pUB110 family, which is predominantly found in Gram-positive bacteria and is known to replicate by the rolling-circle mechanism. The copy number of pTX1 was estimated to be about 150 in each cell. Based on the genetic fingerprints and comparison with other plasmids, it is concluded that pTX1 replicates by a rolling circle mechanism which is rarely found for Pseudomonas plasmids.
We generated gene expression data from the tissues of 50 gastric cancer patients, and applied meta-analysis and gene set analysis to this data and three other stomach cancer gene expression data sets to define the gene expression changes in gastric tumors. By meta-analysis we identified genes consistently changed in gastric carcinomas, while gene set analysis revealed consistently changed biological themes. Genes and gene sets involved in digestion, fatty acid metabolism, and ion transport were consistently down-regulated in gastric carcinomas, while those involved in cellular proliferation, cell cycle, and DNA replication were consistently up-regulated. We also found significant differences between the genes and gene sets expressed in diffuse and intestinal type gastric carcinoma. By gene set analysis of cytogenetic bands, we identified many chromosomal regions with possible gross chromosomal changes (amplifications or deletions). Similar analysis of transcription factor binding sites (TFBSs), revealed transcription factors that may have caused the observed gene expression changes in gastric carcinomas, and we confirmed the overexpression of one of these, E2F1, in many gastric carcinomas by tissue array and immunohistochemistry. We have incorporated the results of our meta- and gene set analyses into a web accessible database (http://human-genome.kribb.re.kr/stomach/).
AL1-gene, necessary for the replication of the genome of a gemini virus TGMV, was inserted in the opposite direction to the promoter CaMV35S resulting in the construction of a plant transformation binary vector pAR35-2. The vector pAR35-2 contains the chimeric gene cassette involving the duplicated promoter CaMV35S, opposite direction of AL1-gene fusioned with hygromycin resistant gene, and the gene cassette of the neomycin phosphotransferase II gene. The plasmid was transferred to tobacco and tomato plants by leaf disk infection via Agrobacterium. The transgenic plants were selected and grown on the MS-agar medium containing kanamycin and hygromycin. The shoots induced from the calli were regenerated to the whole transgenic plants. The antisense AL1-gene was detected in the genomic DNA isolated from the leaves by using the PCR mediated Southern blot analysis. The expression of the antisense AL1-gene was also observed using the RT-PCR mediated Southern blot analysis. The observation of chloroplasts in guard cell pair indicated that the transgenic tomato plants were diploid.
Hepatitis B viral infection which affect about 10% of Korean population manifests asymptomatic carrier, chronic hepatitis and liver cirrhosis and even associates with hepatocellular carcinoma. Clinical manifestations induced by hepatitis B virus vary depending on the degree of immune response by cytotoxic T cells against viral epitope-presenting liver cells. Since hepatitis B virus presents high rate of mutaton that might change the presented epitope and eventually alter immune response, viral mutations, especially in promoters and enhancers, have an important implication in hepatic inflammation and viral replication. To identify mutations related to the hepatic inflammation, we investigated sequence variations of hepatitis B viral promotor regions in the presence or absence of symptoms in hepatitis B carriers. For this, sera from persistently hepatitis B virus-infected mother-child pairs were collected. After PCR amplifiation of all hepatitis B viral promoters (C promoter, S1 promoter, S2/S promoter, X promoter) using serum DNA from each pair, viral promotors were sequenced by automatic sequencer and then sequence data were analyzed by ClustalW. In most cases, the dominant type of maternal virus was transmitted to the child. However, in some children, some new host specific viral variants could be observed in Cp, S1p and S2/Sp. The mutations in C promoter did not seem to be vertically transmitted but arose in new host independently after the wild type had been transmitted. Enhancer I containing X promoter revealed high host specific variations as has been reported before. Two S promoters, S1p and S2/Sp, have shown some point mutations in children, but no deletion mutations were detected as in chronic hepatitis patients in whom deletion mutations are frequently found. In conclusion, the children with the vertically transmitted hepatitis B virus mostly retain the dominant type virus that had been transmitted. However, host specific variants tended to accumulate over time, possibly as clinical symptoms develop.
Kim, Ye-Hwan;Byun, Young Joon;Kim, Won Tae;Jeong, Pildu;Yan, Chunri;Kang, Ho Won;Kim, Yong-June;Lee, Sang-Cheol;Moon, Sung-Kwon;Choi, Yung-Hyun;Yun, Seok Joong;Kim, Wun-Jae
Journal of Korean Medical Science
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v.33
no.47
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pp.303.1-303.10
/
2018
Background: Cell division cycle 6 (CDC6) is an essential regulator of DNA replication and plays important roles in the activation and maintenance of the checkpoint mechanisms in the cell cycle. CDC6 has been associated with oncogenic activities in human cancers; however, the clinical significance of CDC6 in prostate cancer (PCa) remains unclear. Therefore, we investigated whether the CDC6 mRNA expression level is a diagnostic and prognostic marker in PCa. Methods: The study subjects included 121 PCa patients and 66 age-matched benign prostatic hyperplasia (BPH) patients. CDC6 expression was evaluated using real-time polymerase chain reaction and immunohistochemical (IH) staining, and then compared according to the clinicopathological characteristics of PCa. Results: CDC6 mRNA expression was significantly higher in PCa tissues than in BPH control tissues (P = 0.005). In addition, CDC6 expression was significantly higher in patients with elevated prostate-specific antigen (PSA) levels (> 20 ng/mL), a high Gleason score, and advanced stage than in those with low PSA levels, a low Gleason score, and earlier stage, respectively. Multivariate logistic regression analysis showed that high expression of CDC6 was significantly associated with advanced stage (${\geq}T3b$) (odds ratio [OR], 3.005; confidence interval [CI], 1.212-7.450; P = 0.018) and metastasis (OR, 4.192; CI, 1.079-16.286; P = 0.038). Intense IH staining for CDC6 was significantly associated with a high Gleason score and advanced tumor stage including lymph node metastasis stage (linear-by-linear association, P = 0.044 and P = 0.003, respectively). Conclusion: CDC6 expression is associated with aggressive clinicopathological characteristics in PCa. CDC6 may be a potential diagnostic and prognostic marker in PCa patients.
Objective: The present study was executed to explore the molecular mechanism of fibroblast growth factor 10 (FGF10) gene in bovine adipogenesis. Methods: The bovine FGF10 gene was overexpressed through Ad-FGF10 or inhibited through siFGF10 and their negative control (NC) in bovine adipocytes, and the multiplicity of infection, transfection efficiency, interference efficiency were evaluated through quantitative real-time polymerase chain reaction, western blotting and fluorescence microscopy. The lipid droplets, triglycerides (TG) content and the expression levels of adipogenic marker genes were measured during preadipocytes differentiation. The differentially expressed genes were explored through deep RNA sequencing. Results: The highest mRNA level was found in omasum, subcutaneous fat, and intramuscular fat. Moreover, the highest mRNA level was found in adipocytes at day 4 of differentiation. The results of red-oil o staining showed that overexpression (Ad-FGF10) of the FGF10 gene significantly (p<0.05) reduced the lipid droplets and TG content, and their down-regulation (siFGF10) increased the measurement of lipid droplets and TG in differentiated bovine adipocytes. Furthermore, the overexpression of the FGF10 gene down regulated the mRNA levels of adipogenic marker genes such as CCAAT enhancer binding protein alpha (C/EBPα), fatty acid binding protein (FABP4), peroxisome proliferator-activated receptor-γ (PPARγ), lipoprotein lipase (LPL), and Fas cell surface death receptor (FAS), similarly, down-regulation of the FGF10 gene enriched the mRNA levels of C/EBPα, PPARγ, FABP4, and LPL genes (p<0.01). Additionally, the protein levels of PPARγ and FABP4 were reduced (p<0.05) in adipocytes infected with Ad-FGF10 gene and enriched in adipocytes transfected with siFGF10. Moreover, a total of 1,774 differentially expressed genes (DEGs) including 157 up regulated and 1,617 down regulated genes were explored in adipocytes infected with Ad-FGF10 or Ad-NC through deep RNA-sequencing. The top Kyoto encyclopedia of genes and genomes pathways regulated through DEGs were the PPAR signaling pathway, cell cycle, base excision repair, DNA replication, apoptosis, and regulation of lipolysis in adipocytes. Conclusion: Therefore, we can conclude that the FGF10 gene is a negative regulator of bovine adipogenesis and could be used as a candidate gene in marker-assisted selection.
We analyzed QTLs for alkali-related digestion by using 120 population crossed Cheongcheong and Nagdong derived from anther culture (CNDH). The DNA markers located in the QTLs gene were selected and applied to existing cultivars. As a result of the investigation of the alkali decay degree, brown rice of Cheongcheong and Nagdong was 1.9 and 1.6, respectively, and the CNDH was $3.79{\pm}2.01$, and the distribution of variance was distributed to 7.0-1.0. The milled rice of Cheongcheong and Nagdong was 5.6 and 4.1, respectively. The mean of the CNDH was $4.86{\pm}1.55$, and the distribution of variance was distributed to 7.0-2.0. Variation distribution curves showed continuous variation that was close to non-normal distribution. In the QTLs analysis, qBRA2, qBRA6, and qBRA11 were mapped in 1-2 replications of brown rice. QHRA2-1, qHRA2-2, qHRA2-3, qHRA3, and qHRA8 were mapped in the first replication. QHRA2-1, qHRA2-2, qHRA2-3 and qHRA3 were mapped in the second replicates. And mapped to qHRA5 in 4 replicates. These were found on chromosome 2, 3, 6, 8 and 11, respectively. The phenotypic variations of qBRA2, qBRA6, and qBRA11 on the chromosomes of brown and milled rice were 1-9%. The polymorphism was analyzed for 12 types of the japonica type and six types of the indica type, based on the nine markers found in the QTLs analysis of alkali digestion. Chromosome 11, RM27258, was selected to determine the segregation ratio, which shows the difference in size by the band pattern. The results of this study will be used as basic data for the development of high-quality rice cultivars.
An isolate of Cucumber mosaic virus(CMV), called as Can-CMV, was originally isolated from Canna generalis showing typical streak mosaic foliar symptoms, and its properties were investigated in this study. Whereas all known isolates of CMV could induce symptoms on their systemic hosts(four kinds of Nicotiana spp and a zucchini squash), Can-CMV induced no symptoms on its systemic hosts tested. Replication and movement of the virus on upper leaves as well as inoculated leaves-were confirmed by RT-PCR suggesting that Can-CMV could only infect systemically on N. benthamiana and N. glutinosa. Size of local lesions on the Can-CMV-inoculated leaves of Chenopodium amaranticolor was much smaller than that of Fny-CMV. Whereas Fny-CMV and LS-CMV could induce distinct necrotic local lesions on Vigna unguiculata 2 to 3 days postinoculation(dpi), chlorotic spots symptom was expressed by Can-CMV 4 to 5 dpi. Virus-specific 4 kinds of dsRNAs were isolated from leaves of N. benthamiana infected with Can-CMV, and these dsRNAs corresponded to the viral genomic RNAs and subgenomic RNAs and their patterns were indistinguishable to those of Fny-CMV and LS-CMV. By restriction mapping analysis of 950 bp of RT-PCR amplified products of coat protein gene of the virus as well as by serological analysis of gel diffusion test, Can-CMV belongs to a typical member of CMV subgroup IA. These results suggest that the Can-CMV isolated from C. generalis possesses unique pathological properties to understand further insight into the various interactions between virus and host.
Background: Genomic instability, which is manifested by the replication error(RER) phenotype, has been proposed for the promotion of genetic alterations necessary for carcinogenesis. Merlo et al. reported frequent microsatellite instability in primary small cell lung cancers. However, Kim et al. found that instability occurred in only 1% of the loci tested and did not resemble the replication error-positive phenotype. The significance of microsatellite instability in the tumorigenesis of small cell lung cancer as well as the relationship between microsatellite instability and its clinical prognosis was investigated in our study. Methods: Fifteen primary small cell lung cancers were chosen for this study. The DNAs extracted from paraffin-embedded tissue blocks with primary tumor and corresponding control tissue were investigated. Forty microsatellite markers on chromosome 1p, 2p, 3p, 5q, 6p, 6q, 9p, 9q, 13q, and 17p were used in the microsatellite analysis. Results: Thirteen(86.7%) of 15 tumors exhibited LOH in at least one of the tested microsatellite markers. Three of 13 tumors exhibiting LOH lost a larger area in chromosome 9p. LOH was shown in 72.7% on chromosome 2p, 40% on 3p, 50% on 5q, 46.7% on 9p, 69.2% on 13q, and 66.7% on 17p(Table 1). Nine(60%) of 15 tumors exhibited shifted bands in at least one of the tested microsatellite markers. Nine cases exhibiting shifted bands showed altered loci ranging 2.5~52.5%(mean $9.4%\pm16.19$)(Table 2). Shifted bands occurred in 5.7% (34 of 600) of the loci tested(Table 2). Nine cases with shifted bands exhibited LOH ranging between 0~83.3%, and the median survival duration of those cases was 35 weeks. Six cases without shifted bands exhibited LOH ranging between 0~83.3%, and the median survival duration of those cases was 73 weeks. There was no significant difference between median survival durations of the two groups(p=0.4712). Conclusion: Microsatellite instability as well as the inactivation of several tumor suppressor genes may play important roles in the development and progression process of tumors. However, the relationship between microsatellite instability and its clinical prognosis in primary small cell lung cancer could not be established.
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